Hypersensitivity to foods is a world-wide problem in the industrialized world and its prevalence appears to be increasing, affecting approximately 1% of the US population. IgE-mediated reactions to foods, particularly peanuts, are the most common cause of anaphylaxis occurring outside of the hospital. Avoidance of peanuts, the recommended approach, is inadequate due to frequent accidental ingestion. There are a number of emerging experimental therapies but these all have limitations. Although we understand a great deal about how IgE binds to peanut allergens, little is known about how these allergens cross-link IgE/IgE receptor complexes (IgE/FceRI) to activate mast cells and basophils. This proposal will capitalize on three significant findings from our group: 1) the related and complementary 2S albumins, Ara h 2 and Ara h 6, are the most potent peanut allergens, 2) binding of IgE to two specific linear epitopes (one on each allergen) is associated with more severe clinical histories and 3) recombinant Ara h 6 expressed in Pichia pastoris, but not in Escherichia coli, has full effector activity making this a robust model for understanding th molecular basis of effector function. In the proposed studies we will measure binding of patient-derived IgE to specific linear epitopes of Ara h 2 and Ara h 6 in three important clinical settings natural history of clinical reactivity, response of patients to clinically indicated peanut challenes, and response of patients to oral immunotherapy (OIT). We will use these data to determine the relative risk of having defined binding patterns to peptides of Ara h 2 and Ara h 6 in each of these clinical settings. We will then determine the specific amino acids of Ara h 6 and then Ara h 2 that are critical for effective cross-linking of IgE/Fc?RI. Finally we will develop rabbit polyclnal antibodies, murine monoclonal antibodies, and human scFv fragments as potential inhibitors of Ara h 2 and Ara h 6 - mediated cross-linking of IgE/Fc?RI. These will also be useful as novel diagnostic reagents. By completing these specific aims we will overcome the significant obstacle of understanding how allergens interact with IgE to cross-link IgE/FceRI complexes and will move the field forward towards development of new diagnostic and therapeutic tools for the evaluation and treatment of peanut allergy. The future direction of this research is to fully develop these tools and bring them to clinical practice.